B lymphocytes are critical blood cells and altered production or function can cause significant disease. We are interested in understanding how blood stem cells initiate B cell development. Delineating the normal B cell developmental pathway will provide insight into the molecular and cellular basis of many hematologic diseases. Although considerable progress has been made in understanding the generation of naive B cells from committed B cell precursors, little is known about how this process begins. This project will exploit exciting new findings concerning an unexpected regulatory connection between the receptor tyrosine kinase Flt3, HoxA proteins, and the B cell fate determinant EBF. Our new data suggest that this regulatory connection plays an important role in regulating the stem cell to Pro-B transition, and hence, the numbers of B lymphocytes generated. Specifically, we will learn how Flt3 signaling initiates a series of events, gradually biasing multipotential progenitors in bone marrow into the B lineage. Exciting new data revealed that Flt3 signaling, in a ligand-dose dependent fashion, regulates the activity of the primary B cell fate determinant, E2A, a key regulator of the EBF gene. Once expressed, EBF orchestrates B cell differentiation. Gene expression profiling of an EBF-/- cell line pointed to novel role for EBF during B cell development, namely controlling expression of a developmentally regulated cluster of HoxA proteins. Elucidating regulatory circuits that control HoxA expression is important, as dysregulated expression impairs B cell differentiation. First we will determine the molecular events between Flt3 signaling and EBF expression. Then we will determine the regulatory connection between EBF, HoxA, and Flt3 that controls the production of B lymphocytes. Overall, these experiments will provide basic knowledge of a vital process, the production of white blood cells that make antibodies to fight infections. The information should help identify molecular targets for intervention in blood cell cancers and immunodeficiency diseases. PUBLIC HEALTH RELEVANCE: During blood cell development, the stem cell to Pro-B transition is a pivotal checkpoint that controls B cell homeostasis. This project focuses on determining the molecular circuitry through which three critical factors, the receptor tyrosine kinase Flt3, HoxA proteins, and the B cell fate determinant EBF, orchestrate this critical checkpoint.